Further Studies on the Previously Proposed Saturable High Affinity Site for d-Glucose in Renal Brush Border Membrane Preparations

Measurements of the stereospecific uptake of micromolar concentrations of n-glucose by rabbit renal brush border preparations suggested saturation at a "high afanity site" only when incubations at 20" were prolonged, but not when the incubation period was 34 min.This prompted a renewed effort to ascertain whether the apparent saturability at the longer incubation periods was due to bacterial contamination.The following precautionary measures were taken: (a) brush border suspensions were cultured for 24 to 72 hours at 37", and little or no bacterial growth was found; (b) all incubations were carried out in the presence of penicillin and streptomycin; and (c) preincubation of the membrane suspensions at 3'7" decreased the uptake of D- glucose.Although these observations seemed to argue counter to bacterial contamination as a causative factor, subsequent studies revealed the presence in reagent solutions of a temperature (37") sensitive, antibiotic resistant, bacterium of the genus Pseudomonas.These bacteria, isolated from the reagent solutions, cultured at 20", and incubated at 20" with glucose showed a stereospecific uptake of n-glucose, with saturation kinetics at micromolar concentrations and sensitivity to preincubation at 37".With the use of essentially bacteria-free brush border preparations and reagent solutions, saturation was not evident in the high alBnity concentration range of 0.67 to 50 PM D-glucose.It is suggested that there is now no convincing evidence for the presence of the saturable high afllnity site for D-glucose in renal brush borders and that only the Na+-dependent transport of the sugar saturable at higher concentrations is indicated.It is suggested, additionally, that the high aflinity binding of Dglucose to intestinal brush border preparations, reported by others, warrants re-examination.Essentially, sterile renal brush borders bind phloridzin at a high a&rity site, with a &hloridain Of 7 /.cY,Confirming reports of this Site with other renal preparations.